Alternatively spliced exon 5 of the FERM domain of protein 4.1R encodes a novel binding site for erythrocyte p55 and is critical for membrane targeting in epithelial cells.
Direct physical linkage of MAGUKs to the actin cytoskeleton was first established by the interaction of erythrocyte p55 with the FERM domain of protein 4.1R. Subsequently, it was reported that p55 binds to a 51-amino acid peptide, encoded by exon 10, located within the FERM domain of protein 4.1R. In this study, we investigated the nature of the p55-FERM domain binding interface and show that p55 binds to a second 35-amino acid peptide, encoded by an alternatively spliced exon 5, located within the FERM domain of protein 4.1R. Competition and Surface Plasmon Resonance-binding measurements suggest that the peptides encoded by exons 5 and 10 bind to independent sites within the D5 domain of p55. Interestingly, the full length 135 kDa isoform of protein 4.1R containing both exons 5 and 10 was targeted exclusively to the plasma membrane of epithelial cells whereas the same isoform without exon 5 completely lost its membrane localization capacity. Together, these results indicate that p55 binds to two distinct sites within the FERM domain, and the alternatively spliced exon 5 is necessary for the membrane targeting of protein 4.1R in epithelial cells. Since sequences similar to the exon 5-peptide of protein 4.1R and D5 domain of p55 are conserved in many proteins, our findings suggest that a similar mechanism may govern the membrane targeting of other FERM domain containing proteins.
Seo
Pil-Soo
PS
Department of Pharmacology, UIC Cancer Center, University of Illinois College of Medicine, Chicago, IL 60612, USA.
Jeong
Jong-Jin
JJ
Zeng
Lixiao
L
Takoudis
Christos G
CG
Quinn
Brendan J
BJ
Khan
Anwar A
AA
Hanada
Toshihiko
T
Chishti
Athar H
AH
D017398:Alternative Splicing / Q000235:genetics*; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D001798:Blood Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D003598:Cytoskeletal Proteins / Q000737:chemistry* / Q000235:genetics*; D004285:Dogs; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism*; D005091:Exons / Q000235:genetics*; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism*; D008954:Models, Biological; D010455:Peptides / Q000378:metabolism; D011485:Protein Binding; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D021381:Protein Transport
D001798:Blood Proteins; D003598:Cytoskeletal Proteins; C089465:MPP1 protein, human; D008565:Membrane Proteins; D010455:Peptides; D020033:Protein Isoforms; C028841:erythrocyte membrane band 4.1 protein